Epigenetic modifications to DNA and histones mark functionally-important sites in the genome. Assays that identify or 'map'these modifications on a genomewide basis can provide powerful insight into the locations and functions of regulatory elements. However, current epigenomic techniques have limitations, including problems with resolution and an inability to detect allele-specific modifications or repetitive sequences. Bisulphite sequencing is a powerful method for studying DNA methylation, capable of interrogating individual CG dinucleotides. However, this technique does not scale efficiently. Chromatin immunoprecipitation and microarrays can be coupled to achieve global views of histone modifications. However, this technique has suboptimal resolution, is insensitive to repetitive elements and requires large numbers of cells. This project proposes to leverage emerging high-throughput sequencing technologies to overcome the limitations of current tools for epigenome analysis. Our preliminary data demonstrate the applicability of the 454 and Solexa platforms for interrogating epigenetic modifications at unprecedented scale and throughput. The two specific aims of this application propose to apply these instruments to the challenges of mapping cytosine methylation and histone modifications. Aim 1 describes two approaches for analyzing cytosine methylation. In the first, reduced-representation libraries, generated by restriction-digests, will be bisulphite converted and shotgun sequenced to 'read'out their methylation status. In the second, hypomethylated CG dinucleotides will be enriched by affinity capture, and shotgun sequenced. Aim 2 proposes to develop a method that couples chromatin immunoprecipitation and Solexa to map histone modifications genomewide, potentially at higher-resolution, at lower cost, and with lower cell requirements than existing methods. In summary, the proposed projects will harness powerful sequencing technologies and the expertise of a leading genome center to develop new tools for epigenomic analysis that will facilitate the identification of functional genomic elements. As sequencing technology continues to advance, the insights gained from this project will allow their adoption for epigenomic identification of functional elements.